Insight into initiator-DNA interactions: a lesson from the archaeal ORC

Although initiation of DNA replication is considered to be highly coordinated through multiple protein-DNA and protein-protein interactions, it is poorly understood how particular locations within the eukaryotic chromosome are selected as origins of DNA replication. Here, we discuss recent reports that present structural information on the interaction characteristics of the archaeal orthologues of the eukaryotic origin recognition complex with their cognate binding sequences. Since the archaeal replication system is postulated as a simplified version of the one in eukaryotes, by analogy, these works provide insights into the functions of the eukaryotic initiator proteins.     

Anti-stress activity of Propionibacterium freudenreichii: identification of a reactivative protein

Propionibacterium freudenreichii subsp. shermanii is known to prevent mutations caused by various agents such as N-methyl-N'-nitro-N-nitrosoguanidine, 9-aminoacridine, 4-nitro-quinoline-1-oxide and by UV radiation in both prokaryotic and eukaryotic cells. It was also shown to prevent or repair damage caused by H(2)O(2) or UV radiation in Salmonella typhimurium and Escherichia coli, a characteristic previously designated as reactivative effect. In order to characterise this effect at the molecular level, we have purified the active component from a P. freudenreichii cell-free extract using a combination of ammonium sulfate precipitation, anion-exchange and size-exclusion chromatography. The isolated 35 kDa protein was then identified using both N-terminal and internal peptide sequencing as a cysteine synthase. The latter was localised in the P. freudenreichii proteomic map. It is constitutively expressed but also clearly induced during adaptation to detergent and heat, but not acid, stresses. The biological meaning of cysteine synthase in the context of adaptation to oxidative and non-oxidative stresses is discussed.     

Phylogenetic position of Gluconobacter species as a coherent cluster separated from all Acetobacter species on the basis of 16S ribosomal RNA sequences

Abstract The 16S rRNA sequences from the Gluconobacter species G. asaii G. cerinus and G. frateurii were determined and compared with homologous sequences from published databases and sequences of G. oxydans and Acetobacter species previously described [Sievers M., Ludwig W. and Teuber M. (1994) System. Appl. Microbiol. 17, 189–196]. The Gluconobacter species have unique 16S rRNA sequences and exhibit sequence similarity values of 97.4 to 99.1%, corresponding to 36 to 14 base differences. The phylogenetic tree inferring methods (distance matrix, maximum parsimony and maximum likelihood) show that the species of Gluconobacter form a coherent, closely related cluster. Based on the distance matrix method including Rhodopila globiformis as an outgroup reference organism, Gluconobacter is well separated from Acetobacter .     

A novel inhibitor rescues cerebellar defects in a zebrafish model of Down syndrome–associated kinase Dyrk1A overexpression

The highly conserved dual-specificity tyrosine phosphorylation–regulated kinase 1A (Dyrk1A) plays crucial roles during central nervous system development and homeostasis. Furthermore, its hyperactivity is considered responsible for some neurological defects in individuals with Down syndrome. We set out to establish a zebrafish model expressing human Dyrk1A that could be further used to characterize the interaction between Dyrk1A and neurological phenotypes. First, we revealed the prominent expression of dyrk1a homologs in cerebellar neurons in the zebrafish larval and adult brains. Overexpression of human dyrk1a in postmitotic cerebellar Purkinje neurons resulted in a structural misorganization of the Purkinje cells in cerebellar hemispheres and a compaction of this cell population. This impaired Purkinje cell organization was progressive, leading to an age-dependent dispersal of Purkinje neurons throughout the cerebellar molecular layer with larval swim deficits resulting in miscoordination of swimming and reduced exploratory behavior in aged adults. We also found that the structural misorganization of the larval Purkinje cell layer could be rescued by pharmacological treatment with Dyrk1A inhibitors. We further reveal the in vivo efficiency of a novel selective Dyrk1A inhibitor, KuFal194. These findings demonstrate that the zebrafish is a well-suited vertebrate organism to genetically model severe neurological diseases with single cell type specificity. Such models can be used to relate molecular malfunction to cellular deficits, impaired tissue formation, and organismal behavior and can also be used for pharmacological compound testing and validation.     

Adhesion molecules in subjects with COPD and healthy non-smokers: a cross sectional parallel group study

Background

The aim of the study was to investigate how the expression of adhesion molecules changes as neutrophils migrate from the circulation to the lung and if these changes differ between non-smoking subjects and smokers with and without COPD.

Methods

Non-smoking healthy subjects (n=22), smokers without (n=21) and with COPD (n=18) were included. Neutrophils from peripheral blood, sputum and bronchial biopsies were analysed for cell surface expression of adhesion molecules (CD11b, CD62L, CD162). Serum, sputum supernatant and BAL-fluid were analysed for soluble adhesion molecules (ICAM-1, -3, E-selectin, P-selectin, VCAM-1, PECAM-1).

Results

Expression of CD11b was increased on circulating neutrophils from smokers with COPD. It was also increased on sputum neutrophils in both smokers groups, but not in non-smokers, as compared to circulating neutrophils.Serum ICAM-1 was higher in the COPD group compared to the other two groups (p<0.05) and PECAM-1 was lower in smokers without COPD than in non-smoking controls and the COPD group (p<0.05). In BAL-fluid ICAM-1 was lower in the COPD group than in the other groups (p<0.05).

Conclusions

Thus, our data strongly support the involvement of a systemic component in COPD and demonstrate that in smokers neutrophils are activated to a greater extent at the point of transition from the circulation into the lungs than in non-smokers.     

Signaling Governed by G Proteins and cAMP Is Crucial for Growth,Secondary Metabolism and Sexual Development in Fusarium fujikuroi

The plant-pathogenic fungus Fusarium fujikuroi is a notorious rice pathogen causing hyper-elongation of infected plants due to the production of gibberellic acids (GAs). In addition to GAs, F. fujikuroi produces a wide range of other secondary metabolites, such as fusarins, fusaric acid or the red polyketides bikaverins and fusarubins. The recent availability of the fungal genome sequence for this species has revealed the potential of many more putative secondary metabolite gene clusters whose products remain to be identified. However, the complex regulation of secondary metabolism is far from being understood. Here we studied the impact of the heterotrimeric G protein and the cAMP-mediated signaling network, including the regulatory subunits of the cAMP-dependent protein kinase (PKA), to study their effect on colony morphology, sexual development and regulation of bikaverins, fusarubins and GAs. We demonstrated that fusarubin biosynthesis is negatively regulated by at least two Gα subunits, FfG1 and FfG3, which both function as stimulators of the adenylyl cyclase FfAC. Surprisingly, the primary downstream target of the adenylyl cyclase, the PKA, is not involved in the regulation of fusarubins, suggesting that additional, yet unidentified, cAMP-binding protein(s) exist. In contrast, bikaverin biosynthesis is significantly reduced in ffg1 and ffg3 deletion mutants and positively regulated by FfAC and FfPKA1, while GA biosynthesis depends on the active FfAC and FfPKA2 in an FfG1- and FfG3-independent manner. In addition, we provide evidence that G Protein-mediated/cAMP signaling is important for growth in F. fujikuroi because deletion of ffg3, ffac and ffpka1 resulted in impaired growth on minimal and rich media. Finally, sexual crosses of ffg1 mutants showed the importance of a functional FfG1 protein for development of perithecia in the mating strain that carries the MAT1-1 idiomorph.     

Long-Term Effect of Bariatric Surgery on Liver Enzymes in the Swedish Obese Subjects (SOS) Study

Background and Aim

Obesity is associated with elevated serum transaminase levels and non-alcoholic fatty liver disease and weight loss is a recommended therapeutic strategy. Bariatric surgery is effective in obtaining and maintaining weight loss. Aim of the present study was to examine the long-term effects of bariatric surgery on transaminase levels in obese individuals.

Methods

The Swedish Obese Subjects (SOS) study is a prospective controlled intervention study designed to compare the long-term effects of bariatric surgery and usual care in obese subjects. A total of 3,570 obese participants with no excess of alcohol consumption at baseline (1,795 and 1,775 in the control and surgery group, respectively) were included in the analyses. Changes in transaminase levels during follow-up were compared in the surgery and control groups.

Results

Compared to usual care, bariatric surgery was associated with lower serum ALT and AST levels at 2- and 10- year follow up. The reduction in ALT levels was proportional to the degree of weight loss. Both the incidence of and the remission from high transaminase levels were more favorable in the surgery group compared to the control group. Similarly, the prevalence of ALT/AST ratio <1 was lower in the surgery compared to the control group at both 2- and 10-year follow up.

Conclusions

Bariatric surgery results in a sustained reduction in transaminase levels and a long-term benefit in obese individuals.     

External Validation of the Derived Neutrophil to Lymphocyte Ratio as a Prognostic Marker on a Large Cohort of Pancreatic Cancer Patients

Background

With growing evidence on the role of inflammation in cancer biology, the presence of a systemic inflammatory response has been postulated as having prognostic significance in a wide range of cancer types. The derived neutrophil to lymphocyte ratio (dNLR), which represents an easily determinable potential prognostic marker in daily practise and clinical trials, has never been externally validated in pancreatic cancer (PC) patients.

Methods

Data from 474 consecutive PC patients, treated between 2004 and 2012 at a single centre, were evaluated retrospectively. Cancer-specific survival (CSS) was assessed using the Kaplan-Meier method. To evaluate the prognostic relevance of dNLR, univariate and multivariate Cox regression models were applied.

Results

We calculated by ROC analysis a cut-off value of 2.3 for the dNLR to be ideal to discriminate between patients’ survival in the whole cohort. Kaplan-Meier curve reveals a dNLR≥2.3 as a factor for decreased CSS in PC patients (p<0.001, log-rank test). An independent significant association between high dNLR≥2.3 and poor clinical outcome in multivariate analysis (HR = 1.24, CI95% = 1.01–1.51, p = 0.041) was identified.

Conclusion

In the present study we confirmed elevated pre-treatment dNLR as an independent prognostic factor for clinical outcome in PC patients. Our data encourage independent replication in other series and settings of this easily available parameter as well as stratified analysis according to tumor resectability.